Osteopontin (OPN) is a multifunctional, secreted phosphoprotein with beneficial anticalcific effects on pathological calcification. Recent data suggest that OPN may also play a critical role in development of various chronic inflammatory diseases, including atherosclerosis. Furthermore, growing evidence indicates that multiple, proteolytically processed forms of OPN are generated under inflammatory conditions. However, virtually nothing is known about the specific form(s) and function(s) of OPN important in these settings. We propose that a common feature of chronic inflammatory conditions is a requirement for OPN function in macrophages. Specifically, we hypothesize that 1) OPN mediates macrophage recruitment, activation, and survival in atherosclerotic plaques, 2) thrombin, as well as matrix metalloproteases secreted by inflammatory cells modify OPN activity, and 3) that specific OPN proteolytic fragments are required for plaque progression in vivo. Three Specific Aims are proposed to address these hypotheses. In Specific Aim 1, the role of macrophage- and lymphocyte-derived OPN in atherosclerotic lesion initiation and progression in chow-fed ApoE-/- mice will be determined using bone marrow transplant studies. In Specific Aim 2, the role of OPN and its receptors in the modulation of macrophage functions in vitro, including adhesion, migration, survival and activation, will be examined. Finally, in Specific Aim 3, the role of two integrin binding domains, RGD and SLAYGLR domains, of OPN as well as MMPderived OPN proteolytic fragments will be determined during atherosclerotic lesion development and progression in ApoE-/- mice using bone marrow over-expressing these OPN fragments on a macrophage specific promoter. Understanding differences in the mechanisms and structure/function relationships governing inflammatory properties of OPN could help create specific therapeutics targeting these distinct functions.